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At Cain's direction, the Hive was unsealed, releasing zombies and Lickers into the city, and resulting in a catastrophic outbreak of the t-Virus. All efforts to contain the virus failed, and Umbrella sealed off the city in preparation for a nuclear strike. This did not bother Cain in the slightest, as he believed that life was cheap.

It was Cain who woke Alice and Addison, now known as Nemesis, and let them loose in the city, hoping to test them against one another. His plan was undermined by Dr. Charles Ashford, creator of the t-Virus, who enlisted the aid of the survivors in the city to recover his daughter. Although the two subjects engaged each other, there was no clear conclusion to the battle, with Cain finally ordering Nemesis to disengage.

It all came to a head at Raccoon City Hall. In order to convince Alice to fight Nemesis to the death, Cain shot Ashford down in cold blood. When Alice emerged as the victor, Cain tried to convince her to become a part of his plans. When she refused, he ordered Nemesis to kill her.

It was not to be. Matt Addison regained control of his mutated body and the two bioweapons, along with Jill Valentine, Carlos Olivera, and Lloyd Jefferson Wayne began assaulting the Umbrella forces gathered there. Cain then fled to the helicopter, ordering the missile to be fired. He then asks the pilot why the helicopter hadn't taken off yet, to which LJ was revealed to be in the pilot's seat and punches Cain in the face, causing him to fall to the floor. The battle ended with Nemesis being crushed under a helicopter, and the survivors fleeing on a stolen helicopter. Cain tried to bargain for his life, remarking that killing him would not put things right. Alice responded that it wouldn't, but it was a start, and threw him out of the helicopter.

Cain fell to the ground and broke his leg on impact, leaving him incapacitated and at the mercy of the undead, who surrounded and marched towards him. With a nuclear strike moments away, Cain knew he was going to die, but was determined to do so on his own terms. Initially he tried firing a retrieved gun at the approaching zombies, succeeding in killing one, but quickly realized that resistance was pointless. In an attempt to spare himself a painful death at the hands of the zombie horde, Cain put the gun to his head and pulled the trigger, only for it to dry-click, having run out of ammo.

In an act of poetic justice, the first zombie to bite Cain was the last person that he'd killed: Ashford, having reanimated due to either the t-Virus in the air, or the possible t-Virus in his body, meant to restore his handicap like it restored his daughter's. As the zombie horde dogpiled on top of him and slowly ate him alive, Cain came to realize that life was precious, and not cheap, and at the same time, came to see just how cheap he'd allowed his life to become.

The events Cain had set in motion led to the Global T-Virus pandemic; the nuclear strike on Raccoon City did not stop the virus, and it spread throughout the entire planet, drying up lakes and streams, reducing forests to deserts, and infecting humans and animals alike. Within five years, the vast majority of the Earth is little more than a barren wasteland, and only slightly over a million humans remain uninfected. For this, Isaacs regarded Cain as "the man who single-handedly destroyed the world."

Paul W.S. Anderson wrote an alternate ending that was never filmed. The ending involved Cain actually surviving the zombies and nuclear blast by escaping through the sewers leading outside the Raccoon City limits. He would be called and told that Alice had escaped the Umbrella Detroit facility. The scene would then skip to Cain cleaned and talking to Umbrella executives, telling them he will find Alice. He leaves his office and enters his limo, pulls out his laptop and tracks Alice via POV. The POV would show Alice walking towards the limo. She would enter it and kill him while the limo drives away. The ending was scrapped due to the highly unlikely advantages he would make it through the horde of zombies eating him and having time to run into the sewers before the nuclear blast.